Hosoe Misa, Yoshida Nao, Hashiyada Yutaka, Teramoto Hidetoshi, Takahashi Toru, Niimura Sueo
National Institute of Agrobiological Sciences, Tsukuba, Ibaraki 305-8602, Japan.
J Reprod Dev. 2014;60(4):268-73. doi: 10.1262/jrd.2013-110. Epub 2014 Apr 21.
Fetal bovine serum (FBS) has been widely used as a supplement in the maturation medium of bovine oocytes in vitro. However, serum contains many undefined factors and is potentially infectious to humans and animals. As a serum replacement, we evaluated the feasibility of using the silk protein, sericin, derived from the cocoons of silkworm. To examine the rates of oocyte maturation and fertilization, cumulus-oocyte complexes were cultured in TCM-199 supplemented with 0.01%, 0.05%, 0.1% or 0.15% sericin or 5% FBS. The sizes of the perivitelline space that might relate to polyspermy, the expressions of Has2 and CD44 mRNA, the amount of hyaluronan (hyaluronic acid: HA) contained in the oocytes and the rates of blastocyst formation following insemination were then compared between the oocytes cultured with 0.05% sericin and 5% FBS, because the polyspermy rates in oocytes cultured with 0.05% sericin were significantly lower than in those cultured with 5% FBS. After in vitro maturation (IVM), the mean size of the perivitelline space was significantly greater in oocytes cultured with sericin than in those cultured with FBS, although the rates of nuclear maturation, fertilization and blastocyst formation of oocytes under both IVM conditions were not significantly different. The expression of HAS2 and CD44 mRNA and the amount of HA in the denuded oocytes cultured with 0.05% sericin were significantly greater than in those cultured with FBS. These results indicate the feasibility of sericin as an alternative protein supplement for IVM in bovine oocytes.
胎牛血清（FBS）已被广泛用作体外培养牛卵母细胞成熟培养基中的补充剂。然而，血清含有许多未定义的因素，并且对人和动物有潜在的传染性。作为血清替代品，我们评估了使用源自蚕茧的丝蛋白丝胶蛋白的可行性。为了检测卵母细胞成熟和受精率，将卵丘-卵母细胞复合体培养于添加了0.01%、0.05%、0.1%或0.15%丝胶蛋白或5% FBS的TCM-199中。由于用0.05%丝胶蛋白培养的卵母细胞中的多精受精率显著低于用5% FBS培养的卵母细胞，因此随后比较了用0.05%丝胶蛋白和5% FBS培养的卵母细胞之间可能与多精受精相关的卵周隙大小、Has2和CD44 mRNA的表达、卵母细胞中透明质酸（HA）的含量以及授精后囊胚形成率。体外成熟（IVM）后，用丝胶蛋白培养的卵母细胞的卵周隙平均大小显著大于用FBS培养的卵母细胞，尽管在两种IVM条件下卵母细胞的核成熟、受精和囊胚形成率没有显著差异。用0.05%丝胶蛋白培养的裸卵母细胞中HAS2和CD44 mRNA的表达以及HA的含量显著高于用FBS培养的卵母细胞。这些结果表明丝胶蛋白作为牛卵母细胞IVM替代蛋白补充剂的可行性。
